The aspartate pathway of amino acid biosynthesis is one of the central anabolic pathways in bacteria, whose products include four of the eight amino acids that are essential for the nutrition of the adult human. In keeping with its important metabolic function, the aspartate pathway is subject to complex control mechanisms, which attune its activity to the physiological needs of the bacterium. The pattern of control in Bacillus subtilis, the subject of this study, differs fundamentally from that seen in Escherichia coli and is a reflection of the more complex life cycle of B. subtilis, which includes both. a growth and a sporulation phase. The proposed research will focus on the branch of the pathway that leads to the biosynthesis of lysine, in which diaminopimelate is an intermediate as well as an end product in its own right, being a key precursor for the synthesis of the bacterial cell wall peptidoglycan. Earlier work from this laboratory has shown that the control of the reactions leading to diaminopimelate/lysine synthesis in B. subtilis differs from that of all other pathways of amino acid biosynthesis in being under dual control by diaminopimelate and lysine, so as to assure that the pathway can function even in the presence of an excess of the end product lysine or in non- growing cells. The focus of this research is the Control of two operons encoding the key steps of the diaminopimelate/lysine pathway, which have been cloned and sequenced in this laboratory. The dap operon encodes the first three enzymes of diaminopimelate biosynthesis, including the diaminopimelate-sensitive aspartokinase, which catalyzes the rate-determining step in diaminopimelate synthesis, as well as dipicolinate synthase. The lysC operon contains in-phase overlapping genes encoding the subunits of the lysine-sensitive aspartokinase, which catalyzes the rate-limiting step in lysine biosynthesis. Expression of the dap operon probably involves different transcription units and different promoters in growing cells and during sporulation as well as translational attenuation mechanisms. Expression of the lysC operon involves an exceptionally complex transcription control region with a putative transcription attenuator plus possible trans-acting factors. The study of the mechanisms that control the expression of these operons will employ transcript mapping techniques, lacZ gene fusions, site-directed mutagenesis and deletion experiments, analysis of nucleic acid binding proteins, and investigation of the subunit structures of the enzymes aspartokinase I, dihydrodipicolinate synthase, and dipicolinate synthase, which catalyze key steps in the pathway.